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Role of oxidative stress in age-associated mild cognitive impairment and Alzheimer's disease

ROLE OF OXIDATIVE STRESS IN AGE-ASSOCIATED MILD COGNITIVE IMPAIRMENT AND ALZHEIMER’S DISEASE
by
Aaron Clausen
A Dissertation Presented to the
FACULTY OF THE USC GRADUATE SCHOOL
UNIVERSITY OF SOUTHERN CALIFORNIA
In Partial Fulfillment of the
Requirements for the Degree
DOCTOR OF PHILOSOPHY
(NEUROSCIENCE)
May 2011
Copyright 2011 Aaron Clausen

Continuous decline in cognitive performance accompanies the natural aging process in humans, and multiple studies in both humans and animal models have indicated that this decrease in cognitive function is associated with an age-related increase in oxidative stress. Treating aging mammals with exogenous free radical scavengers has generally been shown to attenuate age-related cognitive decline and oxidative stress. I assessed the effectiveness of the superoxide dismutase/catalase mimetics EUK-189 and EUK-207 on age-related decline in cognitive function and increase in oxidative stress. C57/BL6 mice received continuous treatment via osmotic minipumps with either EUK-189 or EUK-207 for 6 months starting at 17 months of age. At the end of treatment, markers for oxidative stress were evaluated by analyzing levels of free radicals, lipid peroxidation and oxidized nucleic acids in brain tissue. In addition, cognitive performance was assessed after 3 and 6 months of treatment with fear conditioning. Both EUK-189 and EUK-207 treatments resulted in significantly decreased lipid peroxidation, nucleic acid oxidation, and reactive oxygen species (ROS) levels. In addition, the treatments also significantly improved age-related decline in performance in the fear-conditioning task. My results thus confirm a critical role for oxidative stress in age-related decline in learning and memory and strongly suggest a potential usefulness for salen–manganese complexes in reversing age-related declines in cognitive function and oxidative load.; A large body of evidence indicates that oxidative stress plays a critical role in normal aging, and in particular in age-related impairment of brain function. Accumulation of oxidatively damaged organelles, proteins, and other macromolecules within various cell types are likely to be involved, as treatment with different types of antioxidants has been shown to reverse some of these effects. Whether cause or consequence, aging is also associated with impairment of the cellular mechanisms involved in removing damaged cellular components, such as the ubiquitin-proteosome and autophagy-lysosome pathways. Furthermore, age-related changes in astrocytes and microglia have also been repeatedly reported. The causal relationships between these multiple age-related changes are far from being clear. I therefore carried out a study directed at testing the role of oxidative stress in age-related changes in autophagy-lysosome function and microglia status. Starting at 17 months of age, C57/BL6 mice received continuous treatment via subcutaneous osmotic pumps with either EUK-189 or EUK-207, two superoxide-dismutase/catalse mimetics that I have previously found to significantly reduce age-dependent oxidative stress and cognitive decline. After 6 months of treatment, markers for autophagy-lysosomal function, and microglia were determined in forebrain. The ratio of LC3-II to LC3-I, which is widely used as a marker of autophagy, was markedly enhanced in brains of 23 month-old mice as compared to that in 16 month-old mice. Expression of the autophagy chaperone protein APG7 decreased with age and lysosomal proteins, such as LAMP-1 and LAMP-2, were significantly reduced in aged brains. These age-related changes in autophagy-lysosme system were reversed by treatment with EUK-189 and EUK-207.; In the CA1 region of the hippocampus, EUK-189 and EUK-207 treatment increased expression of the lysosomal protease cathepsin D, but decreased expression of the autophagy mediator Beclin-1 and lysosomal associated membrane protein LAMP-1. Brain levels of CD11b and numbers of CD11b-imunopositive microglia were decreased in aged brains, suggesting a decline in microglia function. This effect was also reversed by treatment with the SOD/catalase mimetics. Both EUK-189 and EUK-207 have been previously shown to reverse age-related increase in brain oxidative stress, thus these results demonstrate that oxidative stress is causally related to age-related alterations in autophagy-lysosome function and microglia function.; Alzheimer’s disease is an age-associated neurodegenerative disorder that currently affects the lives of about 5.3 million Americans. Alzheimer’s disease (AD) is characterized by progressive memory loss and cognitive deficits that appear to be associated with the accumulation of beta-amyloid plaques and intracellular neurofibrillary tangles within the brain, and neuronal death. A small percentage of AD cases are hereditary and result from mutations in the genes that encode for beta-amyloid precursor protein or proteins responsible for beta-amyloid processing. However, the vast majority of AD cases are sporadic and the underlying molecular mechanisms of AD pathogenesis in these non-hereditary forms remain unclear. A number of studies have shown that, in addition to beta-amyloid and tau pathology, mitochondrial dysfunction and free radical damage are also hallmarks of AD brain, suggesting that oxidative stress might be important in AD pathology. I set out to define the role oxidative stress plays in AD pathogenesis by chronically treating mice that model human AD with the superoxide dismutase (SOD)/catalase mimetic, EUK-207, which I have previously shown to protect against age-associated cognitive impairment and oxidative stress in mice. 3xTg-AD mice, which exhibit cognitive decline as well as Aβ and tau pathology in an age-dependent manner, and wild-type mice were chronically treated with EUK-207 via subcutaneous micro-osmotic pumps for 5 months. Treatment began at 4 months of age because AD pathology and cognitive deficits are still negligible in 3xTg-AD mice at this time point and continued until 9 months of age, when the AD phenotype has been shown to be significant. After 5 months of treatment, cognitive performance was assessed using a fear conditioning paradigm that tests both contextual and cued fear memories. In addition, brain beta-amyloid and tau pathology as well as oxidative stress were analyzed.; At 9 months of age, 3xTg-AD mice exhibited a sharp decline in performance in both contextual and cued fear memory tasks as compared to wild-type mice; however, 3xTg-AD mice administered EUK-207 did not display deficits in fear conditioning performance. Furthermore, chronic treatment with EUK-207 reduced the expression of beta-amyloid, tau and hyperphosphorylated tau accumulation in amygdala and hippocampus of 3xTg-AD mice, and protected against increased levels of Aβ42, oxidized nucleic acids and lipid peroxidation in brain. My results thus confirm a critical role for oxidative stress in AD pathogenesis and strongly suggest a potential usefulness for salen-manganese complexes in protecting against AD development.

ROLE OF OXIDATIVE STRESS IN AGE-ASSOCIATED MILD COGNITIVE IMPAIRMENT AND ALZHEIMER’S DISEASE
by
Aaron Clausen
A Dissertation Presented to the
FACULTY OF THE USC GRADUATE SCHOOL
UNIVERSITY OF SOUTHERN CALIFORNIA
In Partial Fulfillment of the
Requirements for the Degree
DOCTOR OF PHILOSOPHY
(NEUROSCIENCE)
May 2011
Copyright 2011 Aaron Clausen